AU2020281411A1 - Tetracyclic compounds as Cdc7 inhibitors - Google Patents

Abstract

A new class of tetracyclic compounds acting as Cdc7 inhibitors; specifically disclosed are a compound represented by formula (I), isomers thereof, or pharmaceutically acceptable salts thereof.

Description

TETRACYCLIC COMPOUNDS AS CDC7 INHIBITORS CROSS-REFERENCE TO RELATED APPLICATIONS The present application claims the priority to and the benefit of the Chinese Patent Application No. 201910464384.5 filed with China National Intellectual Property Administration on May 30, 2019, the Chinese Patent Application No. 201910491339.9 filed with China National Intellectual Property Administration on Jun. 6, 2019 and the Chinese Patent Application No. 201911128459.9 filed with China National Intellectual Property Administration on Nov. 18, 2019, the disclosure of each of which is incorporated herein by reference in its entirety. TECHNICAL FIELD The present application relates to a new type of tetracyclic compounds as Cdc7 inhibitors, and specifically discloses a compound of formula (I), an isomer thereof or a pharmaceutically acceptable salt thereof. BACKGROUND Cdc7 is a serine/threonine kinase that was first discovered in Saccharomyces cerevisiae in 1974, after which scientists also discovered homologous proteins to it in other eukaryotes. Different species of Cdc7 have certain differences in structure but have very similar functions. In one aspect, it activates MCM to promote the formation of replication origin complexes by phosphorylating minichromosome maintenance proteins (MCM proteins), an important element of DNA replication initiator, and in another aspect, it can also be used as an important regulatory factor of the S-phase checkpoint of a cell cycle for controlling the smooth progress of the cell cycle. HuCdc7, a homologous protein in human cells to Cdc7, was discovered by scientists only in late 1990s. HuCdc7 is expressed in almost all histiocytes of humans. However, it is found that the abnormal high expression of huCdc7 occurs in various tumor cells of humans, and such abnormal high expression shows high correlation with abnormal proliferation and metastasis of tumors and resistance to chemotherapeutic drugs. Therefore, huCdc7 has become an important marker and target in the current tumor research. HuCdc7 is expressed at constant levels in a normal cell cycle and is regulated by several factors and auxiliary proteins in the cell cycle and is therefore in a state of dynamic equilibrium. HuCdc7 is abnormally expressed and over-activated in tumor cells due to disturbances of the cell cycle. Hess et al. found that due to over-expression of huCdc7 in various tumor cells, the over-expressed huCdc7 may promote over-activation of MCM2, an important marker for tumor cells, and thus the abnormal proliferation of tumor cells. Besides, they also found that huCdc7 shows high expression in all metastatic tumor cells, suggesting that the abnormal high expression of huCdc7 may be closely associated with the metastasis of tumor cells. Nambiar et al. have recently found that the auxiliary protein ASK of huCdc7 is also highly expressed in multiple cutaneous melanoma cell lines, which further enhances the activity of huCdc7 in tumor cells. In addition, the abnormal high expression and activation of huCdc7 play a key role in resistance to chemotherapeutic drugs for tumor cells. Tenca et al. found that huCdc7 is extensively expressed with high activity after treating tumor cells with chemotherapeutic drugs Hu and etoposide, and it was noted in the research that huCdc7 inhibits the activity of the two drugs and thus reduces the damage to tumor cells by phosphorylating multiple amino acid sites of MCM2 and MCM4. TAK-931 is a Cdc7 inhibitor and is in phase II clinical trials at present. Therefore, there is a clinical need for developing a new generation of Cdc7 inhibitor capable of being stably metabolized.

HN\ S NH \ N- NN N- N

TAK-931

SUMMARY The present application provides a compound of formula (I), an isomer thereof or a pharmaceutically acceptable salt thereof,

HN\ S NH N L R

(D wherein,

the carbon atom with "*" can be a chiral carbon atom present in a form of a single (R) or (S) enantiomer or in a

form rich in one enantiomer;

L is selected from the group consisting of -CH 2-CH 2-CH 2-, -CH 2-0-CH 2-, -CH 2-S-CH 2-, -CH 2-NH-CH 2-, -NH-CH 2-CH 2-, -S-CH 2-CH 2- and -O-CH2-CH 2-; R 1 is selected from the group consisting of H, halogen, CN, C1 6_ alkyl, C 36_ cycloalkyl, phenyl, and 5-6

membered heteroaryl, wherein the C1-6 alkyl, C 3 _6 cycloalkyl, phenyl, and 5-6 membered heteroaryl are each

independently optionally substituted with 1, 2 or 3 Ra, the 5-6 membered heteroaryl containing 1, 2 or 3

heteroatoms or heteroatom groups each independently selected from the group consisting of 0, S, N and NH;

R2 is selected from Rb, R3 is selected from NH 2, and R 4 is selected from H;

alternatively, R2 is selected from Rc, and R3 and R 4 are joined to form a ring A group optionally substituted

with 1, 2 or 3 Re, wherein the ring A group is selected from the group consisting of C6 _ 14 aryl, 5-14 membered

heteroaryl, 5-12 membered heterocycloalkenyl and 4-14 membered heterocycloalkyl each independently

containing 1, 2 or 3 heteroatoms or heteroatom groups independently selected from the group consisting of 0,

S, N and NRd;

Ra is each independently selected from the group consisting of F, Cl, Br, I, OH, CN, NH 2 , -CH 3 and

Rb is selected from the group consisting of H and C1 6_ alkyl, the Ci-6 alkyl being optionally substituted with 1, 2 or 3 Rbb; Rc is selected from the group consisting of H, F, Cl, Br, I and C1 _3 alkyl;

Rd is selected from the group consisting of H and C1 _4 alkyl;

Rbb is selected from the group consisting of -OCH3 , -OCH 2CH 3, -O-CH(CH 3) 2 , cyclopropyl, cyclopentyl,

phenyl, pyrazolyl, pyridyl, NH 2, -NHCH 3 and -N(CH 3) 2; Re is selected from the group consisting of F, Cl, Br, I, OH, CN, COOH, NH2, -NHCH 3, -N(CH 3)2, -CH 3 ,

-CH 2 CH3 , -CF 3 , -OCH 3 , -OCH2 CH3 , -O-CH(CH 3) 2 , -C(=O)OCH 3 , -C(=O)CH 3 and -C(=O)CH 2 CH3 .

In some embodiments of the present application, the R 1 is selected from the group consisting of H, F, Cl, Br, I, CN, C 1 _3 alkyl, C 3 _5 cycloalkyl, phenyl and 6 membered heteroaryl, wherein the C1 _3 alkyl, C35 cycloalkyl, phenyl, and 6 membered heteroaryl are each independently optionally substituted with 1, 2 or 3 Ra, the 6 membered heteroaryl containing 1, 2 or 3 heteroatoms selected from N. In some embodiments of the present application, the R 1 is selected from the group consisting of H, F, Cl, Br, I, CN, -CH 3, -CH 2CH 3, cyclopropyl, phenyl and pyridyl, wherein the -CH 3, -CH 2CH3, cyclopropyl, phenyl and pyridyl are each independently optionally substituted with 1, 2 or 3 Ra. In some embodiments of the present application, the R 1is selected from the group consisting of H, F, Cl, Br, I, CN, -CH 3, -CH 2CH3, -CF 3, cyclopropyl, phenyl and pyridyl. In some embodiments of the present application, the R1 is selected from H. In some embodiments of the present application, the Ra is selected from F. In some embodiments of the present application, the Rb is selected from the group consisting of H and C1 _4 alkyl, the C 1 _4 alkyl being optionally substituted with 1, 2 or 3 Rbb. In some embodiments of the present application, the Rb is selected from the group consisting of H, methyl, ethyl, isopropyl, n-propyl, n-butyl and isobutyl. In some embodiments of the present application, the Rb is selected from C1 _3 alkyl. In some embodiments of the present application, the Rb is selected from isopropyl. In some embodiments of the present application, the Rc is selected from the group consisting of H, F and C1-3 alkyl. In some embodiments of the present application, the Rc is selected from the group consisting of H, methyl, ethyl and F. In some embodiments of the present application, the Rc is selected from the group consisting of H and methyl. In some embodiments of the present application, the Rd is selected from the group consisting of H, methyl, ethyl, n-propyl, isopropyl and n-butyl. In some embodiments of the present application, the Rd is selected from the group consisting of H and C1-3 alkyl. In some embodiments of the present application, the Rd is selected from the group consisting of H, methyl and isopropyl. In some embodiments of the present application, the ring A group is selected from the group consisting of C 6 _ 10 aryl, 5-9 membered heteroaryl, 5-7 membered heterocycloalkenyl and 4-10 membered heterocycloalkyl optionally substituted with 1, 2 or 3 Re and each independently containing 1, 2 or 3 heteroatoms or heteroatom groups independently selected from the group consisting of 0, S, N and NRd. In some embodiments of the present application, the ring A group is selected from 5-9 membered heterocycloalkyl optionally substituted with 1, 2 or 3 Re, the 5-9 membered heterocycloalkyl containing 1, 2 or 3 heteroatoms or heteroatom groups independently selected from the group consisting of 0, S, N and NRd. In some embodiments of the present application, the ring A group is selected from the group consisting of 5 membered, 6 membered, 7 membered and 8 membered heterocycloalkyl optionally substituted with 1, 2 or 3 Re, the 5 membered, 6 membered, 7 membered and 8 membered heterocycloalkyl containing 1, 2 or 3 heteroatoms or heteroatom groups independently selected from the group consisting of 0, S, N and NRd. In some embodiments of the present application, the ring A group is selected from 5-9 membered heterocycloalkyl optionally substituted with 1, 2 or 3 Re and containing 1 heteroatom or heteroatom group selected from the group consisting of N and NRd.

In some embodiments of the present application, the ring A group is selected from the group consisting of 5 membered, 6 membered, 7 membered and 8 membered heterocycloalkyl optionally substituted with 1, 2 or 3 Re, the 5 membered, 6 membered, 7 membered and 8 membered heterocycloalkyl containing 1 heteroatom or heteroatom group selected from the group consisting of N and NR. In some embodiments of the present application, the ring A group is selected from 5-9 membered heterocycloalkyl containing 1, 2 or 3 heteroatoms or heteroatom groups independently selected from the group consisting of N and NR.

In some embodiments of the present application, the ring A group is selected from 5-9 membered heterocycloalkyl containing 1 heteroatom or heteroatom group selected from the group consisting of N and NRd. In some embodiments of the present application, the ring A group is selected from the group consisting of 5 membered, 6 membered, 7 membered and 8 membered heterocycloalkyl containing 1 heteroatom or heteroatom group selected from the group consisting of N and NR. In some embodiments of the present application, the ring A group is selected from the group consisting of pyrrolidinyl, piperidinyl, morpholinyl, 1-azabicyclo[2.2.2]octyl, 1-azabicyclo[2.2.1]heptanyl, 1-azabicyclo[3.2.2]nonyl and azepanyl optionally substituted with 1, 2 or 3 Re, and contains 1 heteroatom or heteroatom group selected from the group consisting of N and NR. In some embodiments of the present application, the ring A group is selected from the group consisting of pyrrolidinyl, piperidinyl, morpholinyl, 1-azabicyclo[2.2.2]octanyl, 1-azabicyclo[2.2.1]heptanyl, 1-azabicyclo[3.2.2]nonanyl and azepanyl, and contains 1 heteroatom or heteroatom group selected from the group consisting of N and NRd. In some embodiments of the present application, the ring A group is selected

Rd Rd I N N N N

from the group consisting of , , , and optionally substituted with 1, 2 or 3 Re. In some embodiments of the present application, the ring A group is selected from the group consisting of

Rd

1R 3 RR3 In some embodiments of the present application, the structural unit R4 is selected from R Rd Rd R *N N R rN

further from the group consisting of ,,

NRd Rc N Rc

and

R3 In some embodiments of the present application, the structural unit R4 is selected from R4 , the carbon atom to which R3, R4 and Rc are collectively connected being a chiral carbon atom; further from the

R3 R3 I group consisting of R4 and R4 further more from the group consisting of Rc Rd Rd Rd Rd

RN

5 Rd Rd RcNReI Re ReR Re R an N Rc RcV andand Re1RR3RR3

R RC

N KR Rc doRe R N'R Rc R R3 R4$R3 In some embodiments ofthepresent application, thestructural unit R4 is selected from

R Rc rRR

furbonrfromtho pcoRssR4nand Rc ar colctvl cnce bein a*hrlcro tm;frhrfo h

group consistingof R4 and furthermore from the group consisting of R

N RR

dN R RR R

c Rand

R2 -* R, rR 3 R3 In some embodiments of the present application, the structural unit R4 is selected from

H N N .. N .. N/ further from the group consisting of H H N N HNH ii1 ,l 1~ -- ~ 4- NN

and *R2 R R3 R3 In some embodiments of the present application, the structural unit R4 is selected from R4 , the

carbon atom to which R3, R4 and Rc are collectively connected being a chiral carbon atom; further from the

R3 *1 R3 N, group consisting of R4 and R4 further more from the group consisting of

HH H H HHHN N N N NN

C and

R2 x*Rb $R3 R3 In some embodiments of the present application, the structural unit R4 is selected from R4

R NH 2 NH 2 further from Rb , further more from

-,R2 »*Rb R3 -R3 In some embodiments of the present application, the structural unit R4 is selected from R4 , the

carbon atom to which R3, R4 and Rb are collectively connected being a chiral carbon atom; further from the

V Rb Rb SR3 0 R3 NH2 group consisting of R4 and R4 ;further more from the group consisting of Rb and

NH 2 NH 2 2-NH 2 Rb , still further from the group consisting of and In some embodiments of the present application, the L is selected from the group consisting of -CH 2-CH 2-CH 2-, -CH 2-0-CH2-, -CH 2-S-CH 2- and -CH 2-NH-CH 2-. In some embodiments of the present application, the L is selected from -CH 2-CH2-CH 2-. In some embodiments of the present application, the R 1is selected from the group consisting of H, F, Cl, Br, I, CN, -CH 3, -CH 2CH 3, cyclopropyl, phenyl and pyridyl, wherein the -CH 3, -CH 2CH3, cyclopropyl, phenyl and pyridyl are each independently optionally substituted with 1, 2 or 3 Ra, and the other variables are as defined herein. In some embodiments of the present application, the R 1is selected from the group consisting of H, F, Cl, Br, I, CN, -CH 3, -CH 2CH3, -CF 3, cyclopropyl, phenyl and pyridyl, and the other variables are as defined herein. In some embodiments of the present application, the R 1 is selected from H, and the other variables are as defined herein. In some embodiments of the present application, the Rb is selected from the group consisting of H, methyl, ethyl, isopropyl, n-propyl, n-butyl and isobutyl, and the other variables are as defined herein. In some embodiments of the present application, the Rb is selected from isopropyl, and the other variables are as defined herein. In some embodiments of the present application, the Rc is selected from the group consisting of H, methyl, ethyl and F, and the other variables are as defined herein. In some embodiments of the present application, the Rc is selected from the group consisting of H and methyl, and the other variables are as defined herein. In some embodiments of the present application, the Rd is selected from the group consisting of H, methyl, ethyl, propyl, isopropyl and n-butyl, and the other variables are as defined herein. In some embodiments of the present application, the Rd is selected from the group consisting of H, methyl and isopropyl, and the other variables are as defined herein. In some embodiments of the present application, the ring A group is selected from 5-9 membered heterocycloalkyl containing 1 heteroatom or heteroatom group selected from the group consisting of N and NRd, and the other variables are as defined herein. In some embodiments of the present application, the ring A group is selected from the group consisting of

Rd Rd Rd N N N ,f N (7 N and , and the other variables are as defined herein.

*R2 - * R, $FR3 R3 In some embodiments of the present application, the structural unit R4 is selected from R4

R R~Rd Rd R R N c N -d

RRd RcR Rc N'R further from the group consisting of Re

N ,Rc RcK * N

and , and the other variables are as defined herein.

R2 *R i R3 R3 In some embodiments of the present application, the structural unit R4 is selected from R4

H / / NN N N

further from the group consisting of H H I I N H NNNN NNH

NN

and and the other variables are as defined herein.

R2 *Rb R3 $R3 In some embodiments of the present application, the structural unit R4 is selected from R4

NH 2 further from Rb , and the other variables are as defined herein.

R2 5*Rb IR3 R3 In some embodiments of the present application, the structural unit R4 is selected from R4

*NH 2

further from , and the other variables are as defined herein. In some embodiments of the present application, the L is selected from -CH 2-CH 2-CH 2-, and the other variables are as defined herein. In some embodiments of the present application, provided are the compound of formula (I), the isomer thereof or the pharmaceutically acceptable salt thereof, wherein, the carbon atom with "*" can be a chiral carbon atom present in a form of a single (R) or (S) enantiomer or in a form rich in one enantiomer; L is selected from -CH2-CH 2-CH2-; R 1 is selected from H;

R2 is selected from Rb, R3 is selected from NH 2, and R 4 is selected from H;

alternatively, R2 is selected from R, and R3 and R4 are joined to form a ring A group selected from 4-14 membered heterocycloalkyl each independently containing 1, 2 or 3 heteroatoms or heteroatom groups independently selected from the group consisting of N and NRd; Rb is selected from C1 _6 alkyl;

Rc is selected from the group consisting of H and C1-3 alkyl; Rd is selected from the group consisting of H and C1 _4 alkyl.

In some embodiments of the present application, the compound is selected from the group consisting of a compound of formula (I-1) and a compound of formula (1-2), an isomer thereof or a pharmaceutically acceptable salt thereof,

R1 O R,' O HN\ S NH R HN\ S NH R Nb \ 'NN N A N *NH 2

(I-1) (1-2)

wherein, the carbon atom with "*" can be a chiral carbon atom present in a form of a single (R) or (S) enantiomer or in a form rich in one enantiomer; R 1 and Rc are as defined in the compound of formula (I) disclosed herein; the ring A group is as defined in the compound of formula (I) disclosed herein; Rb is as defined for the compound of formula (I) disclosed herein.

*A In some embodiments of the present application, the structural unit is defined as the structural unit

R4 R3 is.

In some embodiments of the present application, the compound is selected from the group consisting of a compound of formula (I-la), a compound of formula (1-lb), a compound of formula (I-2a) and a compound of formula (I-2b), an isomer thereof or a pharmaceutically acceptable salt thereof, R1 O R1' O

HN NHR H HN N N SK

Rc HN S NH NN" \ N Rb \

NH 2

(I-2b)

wherein,

R 1 and Rc are as defined in the compound of formula (I) disclosed herein, and the carbon atom to which Rc and the ring A group are collectively connected is a chiral carbon atom; the ring A group is as defined in the compound of formula (I) disclosed herein; Rb is as defined for the compound of formula (I)disclosed herein.

AA In some embodiments of the present application, the structural unit or is defined as

6 R3 ~SR3 the structural unit R4 or R4 is, respectively. In some embodiments of the present application, the compound is selected from the group consisting of a compound of formula (I-11), a compound of formula (1-12), a compound of formula (1-13), a compound of formula (1-14), a compound of formula (1-15) and a compound of formula (1-16), an isomer thereof or a pharmaceutically acceptable salt thereof,

HN S NH R HN\ S NH R HN S NH R Ns \ |c N:\:N N N NRd N N N NRd

(I-11) (1-12) (1-13) 0 o 0 HN S NH RN\ S NH R H S NN\ / N N\ RN N N

N~d(1-15) N (1-14) Rd (1-16)

wherein, the carbon atom with "*" is a chiral carbon atom present in a form of a single (R) or (S) enantiomer or in a form rich in one enantiomer; R1, Rc and Rd are as defined for the compound of formula (I)disclosed herein. In some embodiments of the present application, the compound is selected from the group consisting of a compound of formula (I-1la), a compound of formula (I-1lb), a compound of formula (I-12a), a compound of formula (I-12b), a compound of formula (I-13a), a compound of formula (I-13b), a compound of formula (I-14a), a compound of formula (I-14b), a compound of formula (I-16a) and a compound of formula (I-16b), an isomer thereof or a pharmaceutically acceptable salt thereof,

H\ S-NHRHR 0H

HN NRd NIRd H

H\ S HN\ N-- ( I-11a) NNH S NHN ( I-11b) sH( I-12a) (I-12b) H \\ N-RH

d NNRd

N NR NNNR H

(I-13a) (I-13b) (I-14a)

N) 0 0 N~Rd

HN S NH HNR H N R

(I-14b) (I-16a) (I-16b)

wherein, R 1, Rcand Rdare as defined for the compound of formula (I) disclosed herein. 100 In some aspects, the present application encompasses the variables defined above and solutions thereof, as well as any combination thereof. HN- \ NH R. N/-\ N N \ rj-. \ The present application also provides a compound of the formula below, an isomer thereof or a pharmaceutically acceptable salt thereof, N N 0oN N 0 NH N

N NH, eie o h N, opudo oml 1 icoe Nan[ een Ndaea HN\ NH HN S NH HN S NH HN NH

N N N N N

0 o 0 0

NH HN N N HN NH HN \ S NH2 HN\ 1

0 o 0 0 HN\ SHS S HHJ ~N HNJ S1 N

Insomeembodiments ofthepresent application, provided arethe compound, the isomer thereof or the ANN N N- N NN N N"N pharmaceutically acceptable salt thereof,selectedfromthegroupconsistingof

HN H\ NH HN\ S NHl HN NH

H N o o 0 0 HN\ S NH HN\& S NH HN NH HN\ S NH N N\ 'N N N o o 0 0 H N H NH HIN H NH N-H N N \N 1.N HN\ S NHN S NH H N H o 0 0 0

HN S N HN S 1 NH HN S NH HN S NH N N NN

The present application further provides a pharmaceutical composition comprising a therapeutically effective

amount of the compound, the isomer thereof or the pharmaceutically acceptable salt thereof and a

pharmaceutically acceptable carrier.

The present application also provides use of the compound, the isomer thereof or the pharmaceutically

acceptable salt thereof or the pharmaceutical composition in preparing a therapeutic Cdc7 inhibitor.

The present application also provides use of the compound, the isomer thereof or the pharmaceutically

acceptable salt thereof or the pharmaceutical composition in preparing a medicament for treating a tumor.

The present application also provides a method for treating a Cdc7 kinase-mediated disease, comprising

administering to a mammal, preferably a human, in need of such treatment a therapeutically effective amount

of the compound, the isomer thereof or the pharmaceutically acceptable salt thereof or the pharmaceutical

composition.

The present application also provides use of the compound, the isomer thereof or the pharmaceutically

acceptable salt thereof or the pharmaceutical composition in treating a Cdc7 kinase-mediated disease.

The present application also provides the compound, the isomer thereof or the pharmaceutically acceptable salt

thereof or the pharmaceutical composition for use in treating a Cdc7 kinase-mediated disease.

The present application also provides the compound, the isomer thereof or the pharmaceutically acceptable salt

thereof for use as a medicament. In some embodiments of the present application, the use as a medicament

refers to use as a medicament for treating a Cdc7 kinase-mediated disease.

In some embodiments of the present application, the Cdc7 inhibitor is a medicament for treating a tumor.

In some embodiments of the present application, the tumor includes colorectal cancer and pancreatic cancer.

In some embodiments of the present application, the medicament for treating a tumor refers to a medicament

for treating colorectal cancer and pancreatic cancer.

Technical Effects As a Cdc7 inhibitor, the compound disclosed herein has a wide application prospect in treating tumors. The

compound disclosed herein has strong inhibitory activity against Cdc7/DBF4, and also shows good inhibitory

activity against Colo205 cells. In addition, the compound disclosed herein has good AUCoat and

bioavailability and significant inhibitory effects on tumors in mice. Therefore, further intensive research on the

Cdc7 kinase and inhibitors thereof is expected to pave a new way for clinically treating tumors. The compound disclosed herein is expected to become a new medicament with better therapeutic effects and lower toxic and side effects compared to similar products. Definitions and Description Unless otherwise stated, the following terms and phrases used herein are intended to have the following meanings. A particular term or phrase, unless otherwise specifically defined, should not be considered as uncertain or unclear, but construed according to its common meaning. When referring to a trade name, it is intended to refer to its corresponding commercial product or its active ingredient. The term "pharmaceutically acceptable" is used herein for those compounds, materials, compositions and/or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problems or complications, and commensurate with a reasonable benefit/risk ratio. The term "pharmaceutically acceptable salt" refers to a salt of the compound disclosed herein, which is prepared from the compound having particular substituents disclosed herein and a relatively nontoxic acid or base. When the compound disclosed herein contains a relatively acidic functional group, a base addition salt can be obtained by contacting such a compound with a sufficient amount of a base in a pure solution or a suitable inert solvent. Pharmaceutically acceptable base addition salts include sodium, potassium, calcium, ammonium, organic amine, or magnesium salts, or similar salts. When the compound disclosed herein contains a relatively basic functional group, an acid addition salt can be obtained by contacting such a compound with a sufficient amount of an acid in a pure solution or a suitable inert solvent. Examples of pharmaceutically acceptable acid addition salts include salts derived from inorganic acids, such as hydrochloric acid, hydrobromic acid, nitric acid, carbonic acid, bicarbonate radical, phosphoric acid, monohydrogen phosphate, dihydrogen phosphate, sulfuric acid, hydrogen sulfate, hydroiodic acid and phosphorous acid; and salts derived from organic acids, such as acetic acid, propionic acid, isobutyric acid, maleic acid, malonic acid, benzoic acid, succinic acid, suberic acid, fumaric acid, lactic acid, mandelic acid, phthalic acid, benzenesulfonic acid, p-toluenesulfonic acid, citric acid, tartaric acid and methanesulfonic acid. Also included are salts of amino acids (e.g., arginine) and salts of organic acids such as glucuronic acid. Some particular compounds disclosed herein contain both basic and acidic groups and thus can be converted to any base or acid addition salts. The pharmaceutically acceptable salts disclosed herein can be synthesized from a parent compound having an acidic or basic group by conventional chemical methods. In general, such salts are prepared by the following method: the free acid or base form of the compound reacting with a stoichiometric amount of the appropriate base or acid in water or an organic solvent or a mixture thereof. The pharmaceutically acceptable salt disclosed herein can be converted into the free state by using a known method or a method similar thereto, for example, by reacting a pharmaceutically acceptable acid addition salt or base addition salt with a stoichiometric amount of an appropriate base or acid. The compounds disclosed herein can be in the form of a geometric isomer or stereoisomer. All such compounds are contemplated herein, including cis and trans isomers, (-)- and (+)- enantiomers, (R)- and (S) enantiomers, diastereoisomers, (D)-isomers, (L)-isomers, and racemic mixtures and other mixtures thereof, such as an enantiomer or diastereoisomer enriched mixture, all of which are encompassed within the scope of the present application. Substituents such as alkyl may have an additional asymmetric carbon atom. All these isomers and mixtures thereof are encompassed within the scope of the present application. The carbon atom with "*" disclosed herein may be a chiral carbon atom, which means either a chiral carbon atom or an achiral carbon atom, depending on the connection of the carbon atom in the structure of a compound. Unless otherwise stated, the term "enantiomer" or "optical isomer" refers to stereoisomers that are mirror images of each other. Unless otherwise stated, the term "cis-transisomer" or "geometric isomer" results from the inability of a single bond of a ring carbon atom or a double bond to rotate freely. Unless otherwise stated, the term "diastereomer" refers to stereoisomers whose molecules have two or more chiral centers and are not mirror images of each other. Unless otherwise stated, "(+)" stands for dextrorotation, "(-)" stands for levorotation, and "(±)" stands for racemization. Unless otherwise stated, the absolute configuration of a stereogenic center is represented by a wedged solid bond ( ') and a wedged dashed bond (.- `), and the relative configuration of a stereogenic center is represented by a straight solid bond ( /) and a straight dashed bond (,-'). A wavy line ( -) represents a wedged solid bond ( 4) or a wedged dashed bond ('),or a wavy line ( )represents a straight solid bond ( /) or a straight dashed bond ( ). Unless otherwise stated, the term "rich in one isomer", "isomer-rich", "rich in one enantiomer", or "enantiomer-rich" means that the content of one of the isomers or enantiomers is less than 100% and more than or equal to 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, 99%, 99.5%, 99.6%, 99.7%, 99.8% or 99.9%. Optically active (R)- and (S)-isomers and D and L isomers can be prepared by chiral synthesis or chiral reagents or other conventional techniques. An enantiomer of certain compound disclosed herein can be prepared by asymmetric synthesis or derivatization using a chiral auxiliary, wherein the resulting diastereoisomeric mixture is separated and the auxiliary group is cleaved so as to provide the desired pure enantiomer. Alternatively, when the molecule contains a basic functional group (such as amino) or an acidic functional group (such as carboxyl), the compound reacts with an appropriate optically active acid or base to form a salt of the diastereoisomer, which is then subjected to diastereoisomeric resolution through conventional methods in the art to give the pure enantiomer. Furthermore, the enantiomer and the diastereoisomer are generally isolated through chromatography using a chiral stationary phase, optionally in combination with chemical derivatization (e.g., carbamate generated from amines). The compound disclosed herein may contain an unnatural proportion of atomic isotope at one or more of the atoms that constitute the compound. For example, the compound may be labeled with a radioisotope, such as tritium 3( H), iodine-125

( 125I), or C-14 (4C). For another example, hydrogen can be substituted with deuterium to form a deuterated drug, and the bond formed by deuterium and carbon is firmer than that formed by common hydrogen and carbon. Compared with an un-deuterated drug, the deuterated drug has the advantages of reduced toxic side effect, increased stability, enhanced efficacy, prolonged biological half-life and the like. All isotopic variations of the compound described herein, whether radioactive or not, are encompassed within the scope of the present application.

Unless otherwise specified, the compounds disclosed herein include both E and Z geometric isomers when they contain olefinic double bonds or other centers of geometric asymmetry. Likewise, all tautomeric forms R1 R HN\ N N HN are encompassed within the scope of the present application. For example, and are

NH OH

tautomeric to each other; for another example, ' N , H and N' are

tautomeric to one another. The term "substituted" means that one or more hydrogen atoms on a specific atom are substituted with substituents which may include deuterium and hydrogen variants, as long as the valence of the specific atom is normal and the substituted compound is stable. When the substituent is an oxygen (i.e., =0), it means that two hydrogen atoms are substituted. Substitution of oxygen does not occur on aromatic groups. The term "optionally substituted" means that an atom can be substituted with a substituent or not. Unless otherwise specified, the type and number of the substituent may be arbitrary as long as being chemically achievable. When any variable (e.g., R) occurs more than once in the constitution or structure of a compound, the variable is independently defined in each case. Thus, for example, if a group is substituted with 0-2 R, the group can be optionally substituted with two R at most, and the definition of R in each case is independent. Furthermore, a combination of a substituent and/or a variant thereof is permissible only if the combination can result in a stable compound. When a bond of a substituent can be connected to one or more atoms on a ring, the substituent can be bonded

to any atom on the ring. For example, the structural unit - or represents that the substitution of substituent R may occur in any one position on cyclohexyl or cyclohexadienyl. When it is not specified by which atom the listed substituent is connected to the group to be substituted, the substituent can be connected via any atom of the group. For example, pyridinyl as a substituent can be connected to the group to be substituted through any carbon atom on the pyridine ring. When bonds of two substituents can be connected to the same carbon atom on a ring, the substituents can be H N

bonded to any one of the same carbon atoms on the ring. For example, the structural unit represents that the substitution of two substituents may occur at any one of the same carbon atoms of the piperidine ring, H H N H H N NoN N

and thus, the structural unit includes , and ' , but not structural units

H H N H H N sNN

such as ,I- or . When the enumerative linking group does not

indicate the direction for linking, the direction for linking is arbitrary. For example, when the linking group L

A L B contained in is -M-W-, -M-W- can either link ring A and ring B in a direction same as

left-to-right reading order to form M-W , or link ring A and ring B in an opposing

A W-M B direction to form . A combination of the linking group, a substituent and/or a variant thereof is permissible only if the combination can result in a stable compound. Unless otherwise specified, when a group has one or more connectable sites, any one or more of the sites of the group may be connected to other groups by chemical bonds. When there is no designated connecting mode for a chemical bond and H atoms are present at a connectable site, the number of the H atoms at the connectable site is correspondingly reduced based on the number of the connected chemical bonds, and a group with a corresponding valence number is thus formed. The chemical bond that connects the site to another group may

be represented by a straight solid bond ( straightgt dashed bond( ), or a wavy line ( For

example, the straight solid bond in -OCH3 refers to being connected to another group via the oxygen atom in

the group; the straight dashed bond in H refers to being connected to another group via two ends of the

nitrogen atom in the group; the wavy line in refers to being connected to another group via the

NH carbon atoms at positions 1 and 2 in the phenyl group; I means that any connectable site on the

piperidinyl can be connected to another group via 1 bond, and at least 4 connecting modes 7 -,

NH NH CNH -- NH | , and are possible; even if -N- is connected to an H atom, I

includes the connecting mode of , except that when 1 bond is connected to a site, the number of H at that site is correspondingly reduced by 1 and a monovalent piperidinyl is thus formed.

Unless otherwise specified, the number of atoms on a ring is generally defined as the member number of the ring. For example, "5-7 membered ring" refers to a "ring" on which 5 to 7 atoms are arranged in a circle.

Unless otherwise specified, the term "Ci-6 alkyl" refers to a linear or branched saturated hydrocarbon group consisting of 1 to 6 carbon atoms. The Ci-6 alkyl includes C1 _5 , Ci_ 4 , C1 _ 3 , C1 _ 2 , C2 6_ , C 2 _4, C6 , and C5 alkyl,

etc., and may be monovalent (e.g., methyl), divalent (e.g., methylene), or polyvalent (e.g., methenyl). Examples of C 1 _ 6 alkyl include, but are not limited to, methyl (Me), ethyl (Et), propyl (including n-propyl and isopropyl), butyl (including n-butyl, isobutyl, s-butyl, and t-butyl), pentyl (including n-pentyl, isopentyl, and neopentyl), hexyl, and the like. Unless otherwise specified, the term "halogen", by itself or as part of another substituent, refers to a fluorine, chlorine, bromine or iodine atom. Unless otherwise specified, "C3_ 6 cycloalkyl" refers to a saturated cyclic hydrocarbon group consisting of 3 to 6 carbon atoms, which is a monocyclic and bicyclic ring system, wherein the carbon atoms may optionally be oxidized (i.e., C=O). The C 3 _ 6 cycloalkyl includes C 3 _ 5, C 4 _ 5, C5 6 cycloalkyl and the like, and may be monovalent, divalent or polyvalent. Examples of C 3 _6 cycloalkyl include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl and the like. Unless otherwise specified, the term "4-14 membered heterocycloalkyl", by itself or in combination with other terms, refers to a saturated cyclic group consisting of 4 to 14 ring atoms, of which 1, 2, 3 or 4 ring atoms are heteroatoms independently selected from the group consisting of 0, S, and N, with the remaining being carbon atoms. The nitrogen atom is optionally quaternized, and the carbon, nitrogen and sulfur heteroatoms can be optionally oxidized (i.e., C=, NO and S(O)p, wherein p is 1 or 2). This includes monocyclic, bicyclic and tricyclic systems, wherein the bicyclic and tricyclic systems include spirocyclic, fused and bridged rings. Furthermore, with respect to the "4-14 membered heterocycloalkyl", a heteroatom may occupy the position where the heterocycloalkyl is connected to the rest of the molecule. The 4-14 membered heterocycloalkyl includes 4-12 membered, 4-10 membered, 5-10 membered, 5-9 membered, 5-8 membered, 3-10 membered, 3-8 membered, 3-6 membered, 3-5 membered, 4-6 membered, 5-6 membered, 4 membered, 5 membered and 6 membered heterocycloalkyl and the like. Examples of 4-14 membered heterocycloalkyl include, but are not limited to, azetidinyl, oxetanyl, thietanyl, pyrrolidinyl, pyrazolidinyl, imidazolidinyl, tetrahydrothienyl (including tetrahydrothien-2-yl, tetrahydrothien-3-yl, etc.), tetrahydrofuranyl (including tetrahydrofuran-2-yl, etc.), tetrahydropyranyl, piperidinyl (including 1-piperidinyl, 2-piperidinyl, 3-piperidinyl, etc.), piperazinyl (including 1-piperazinyl, 2-piperazinyl, etc.), morpholinyl (including 3-morpholinyl, 4-morpholinyl, etc.), dioxanyl, dithianyl, isoxazolidinyl, isothiazolidinyl, 1,2-oxazinyl, 1,2-thiazinyl, hexahydropyridazinyl, N NH NH

homopiperazinyl, homopiperidinyl, dioxepanyl, , or the like. Unless otherwise specified, the term "5-12 membered heterocycloalkenyl", by itself or in combination with other terms, refers to a partially unsaturated cyclic group consisting of 5 to 12 ring atoms containing at least one carbon-carbon double bond, of which 1, 2, 3 or 4 ring atoms are heteroatoms independently selected from the group consisting of 0, S, and N, with the remaining being carbon atoms. The nitrogen atom is optionally quaternized, and the carbon, nitrogen and sulfur heteroatoms can be optionally oxidized (i.e., C=O, NO and S(O)p, wherein p is 1 or 2). This includes monocyclic, bicyclic and tricyclic systems, wherein the bicyclic and tricyclic systems include spirocyclic, fused and bridged rings, and any ring of these systems is nonaromatic. Furthermore, with respect to the "5-12 membered heterocycloalkenyl", a heteroatom may occupy the position where the heterocycloalkenyl is connected to the rest of the molecule. The 5-12 membered heterocycloalkenyl includes 5-10 membered, 5-8 membered, 5-6 membered, 4-5 membered, 4 membered, 5 membered, 6 membered heterocycloalkenyl and the like. Examples of 5-12 membered heterocycloalkenyl include, but are not limited to, 0 , 47,N S, N7>N aON H , NH , N NO or H 0

Unless otherwise specified, the term "C 6 _ 14 aryl" herein refers to a cyclic hydrocarbon group consisting of 6 to 14 carbon atoms and having a conjugated 7-electron system, which may be a monocyclic, fused bicyclic, or fused tricyclic ring system, wherein each ring is aromatic. It may be monovalent, divalent or polyvalent, and the C6 _ 14 aryl includes C6 _ 10 , C6 _ 9, C6 -, C1 2 , C 14 , CIO and C 6 aryl and the like. Examples of C6 _ 14 aryl include,

but are not limited to, phenyl, naphthyl (including 1-naphthyl, 2-naphthyl, etc.) and anthryl. Unless otherwise specified, the term "5-14 membered heteroaryl" herein refers to a cyclic group consisting of 5 to 14 ring atoms and having a conjugated 7-electron system, of which 1, 2, 3 or 4 ring atoms are heteroatoms independently selected from the group consisting of 0, S and N, with the remaining being carbon atoms. The nitrogen atom is optionally quaternized, and the carbon, nitrogen and sulfur heteroatoms may optionally be oxidized (i.e., C=, NO and S(O)p, wherein p is 1 or 2). It can be a monocyclic, fused bicyclic or fused tricyclic system, wherein the rings are aromatic. The 5-14 membered heteroaryl can be linked to the rest of the molecule through a heteroatom or a carbon atom. The 5-14 membered heteroaryl includes 5-12 membered, 5-10 membered, 5-8 membered, 5-7 membered, 5-6 membered, 5 membered, 6 membered heteroaryl and the like. Examples of the 5-12 membered heteroaryl include, but are not limited to, pyrrolyl (including N-pyrrolyl, 2-pyrrolyl, 3-pyrrolyl, etc.), pyrazolyl (including 2-pyrazolyl, 3-pyrazolyl, etc.), imidazolyl (including N-imidazolyl, 2-imidazolyl, 4-imidazolyl, 5-imidazolyl, etc.), oxazolyl (including 2-oxazolyl, 4-oxazolyl, 5-oxazolyl, etc.), triazolyl (including 1H-1,2,3-triazolyl, 2H-1,2,3-triazolyl, 1H-1,2,4-triazolyl, 4H-1,2,4-triazolyl, etc.), tetrazolyl, isoxazolyl (including 3-isoxazolyl, 4-isoxazolyl, 5-isoxazolyl, etc.), thiazolyl (including 2-thiazolyl, 4-thiazolyl, 5-thiazolyl, etc.), furanyl (including 2-furanyl, 3-furanyl, etc.), thienyl (including 2-thienyl, 3-thienyl, etc.), pyridinyl (including 2-pyridinyl, 3-pyridinyl, 4-pyridinyl, etc.), pyrazinyl, pyrimidinyl (including 2-pyrimidinyl, 4-pyrimidinyl, etc.), benzothiazolyl (including 5-benzothiazolyl, etc.), purinyl, benzimidazolyl (including 2-benzimidazolyl, etc.), benzoxazolyl, indolyl (including 5-indolyl, etc.), isoquinolinyl (including 1-isoquinolinyl, 5-isoquinolinyl, etc.), quinoxalinyl (including 2-quinoxalinyl, 5-quinoxalinyl, etc.), quinolyl (including 3-quinolyl, 6-quinolyl, etc.) or benzoisoquinolin. Unless otherwise specified, the term "heteroatom or heteroatom group (i.e., heteroatom-containing atom group)" includes atoms other than carbon (C) and hydrogen (H) as well as atom groups containing these heteroatoms, for example, oxygen (0), nitrogen (N), sulfur (S), silicon (Si), germanium (Ge), aluminum (Al), boron (B), -0-, -S-, =0, =S, -C(=0)O-, -C(=)-, -C(=S)-, -S(=), -S(=0)2-, and optionally substituted -C(=0)N(H)-, -N(H)-, -C(=NH)-, -S(=0)2 N(H)- or -S(=0)N(H)-. The compounds disclosed herein can be prepared by a variety of synthetic methods well known to those skilled in the art, including the specific embodiments listed below, embodiments formed by combinations thereof with other chemical synthetic methods, and equivalents thereof known to those skilled in the art. The preferred embodiments include, but are not limited to, the examples disclosed herein.

An important consideration in synthesis route planning in the art is the selection of suitable protecting groups for reactive functional groups (e.g., amino in the present application). For example, reference may be made to Greene's Protective Groups in Organic Synthesis (4th Ed.) Hoboken, New Jersey: John Wiley & Sons, Inc. All references cited herein are incorporated by reference in their entirety. General synthetic route:

N-.

0 NH2 HN CN C 0 N 2

CN N

0 0 R_ .2N 1R3 - S \I NH 2 _ _ _ _ _KA 0

NH NS NH R2 N- R2

0 0 R4 R4 1.Deprotection HN \ S + HN\ S NH 2, SFC N- N' 4.R2 N- N R2

wherein,

N/\ R'is Cl or 0 R2, R3 and R 4 are as described for the compound of formula (I). The solvents used herein can be commercially available. The following abbreviations are used herein: DMF represents N,N-dimethylformamide; DMSO represents dimethyl sulfoxide; BID represents administration twice daily. Compounds are named according to conventional nomenclature rules in the art or using ChemDraw@ software, and supplier's catalog names are given for commercially available compounds. BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 is a graph of tumor growth curves of tumor-bearing mice of a xenograft tumor model of the human colorectal cancer cell SW620 after administration of test compounds; and FIG. 2 is a picture of tumors of mice in a subcutaneous xenograft tumor nude mouse model of the human colorectal cancer cell SW620. DETAILED DESCRIPTION The present application is described in detail below by way of examples. However, this is by no means disadvantageously limiting the scope of the present application. Although the present application has been described in detail herein and specific examples have also been disclosed, it will be apparent to those skilled in the art that various changes and modifications can be made to the specific embodiments without departing from the spirit and scope of the present application. Example 1. Compound 1-1 Hydrochloride and Compound 1-2 Hydrochloride

NH

0 HN

1A 1B 1C

O HS NH 2 NH HN _ 1iF10 N NH NCI 0_____

HN CN CN O 2 NH 20 1D 1E 1G

0 o 0 CH N \ S NH 2 0 O N NH N S NH

o N

11 1J

0 0

HN\ S NH - HN S NH N N- N NN/

1-1 1-2

Preparation of compound IB: Compound 1A (2 g) was stirred in N,N-dimethylformamide dimethyl acetal (5.67 g) at 100 °C for 3 h. After the reaction was completed, the reaction mixture was concentrated to dryness under reduced pressure to give compound 1B for direct use in the next step without purification. LCMS (ESI) m/z: 182 (M+1). Preparation of compound IC: Hydrazine hydrate (832.40 mg) was added to a solution of 1B (2.87 g) in methanol (6 mL) at 0 °C, followed by heating to 90 °C and stirring for 2 h. After the reaction was completed, the reaction mixture was concentrated to dryness under reduced pressure, followed by addition of 5 mL of a mixed solution (ethyl acetate/petroleum ether = 1/10, v/v), and the resultant mixture was stirred for 5 min and then filtered to give solid compound IC. LCMS (ESI) m/z: 151 (M+1). Preparation of compound ID: DMF (987.06 mg) was slowly dropwise added to phosphorus oxychloride (2.07 g) at 0 °C, followed by stirring for 10 min, with white solid precipitated. Compound IC (1.2 g) was dissolved in phosphorus oxychloride (30 mL), and the resultant solution was added to the above solid. The reaction mixture was then warmed to 50 °C, added with hydroxylamine hydrochloride (1.67 g) and stirred at 50 °C for 2 h. After the reaction was completed, the reaction mixture was distilled under reduced pressure to remove excess phosphorus oxychloride, and the residue was dissolved in DMF and purified by reversed-phase flash chromatography

(Agilent, C18 reversed-phase column, 20-35 [tm, 0.1% aqueous formic acid solution/acetonitrile) to give compound ID. LCMS(ESI)m/z: 194(M+1); 'HNMR(METHANOL-d4) 6 ppm 7.87-7.92(m, 1H), 3.02-3.06(m, 2H), 2.70-2.78(m, 2H), 1.99-2.07(m, 2H). Preparation of compound 1E: Compound ID (1.6 g), 3,4-dihydro-2H-pyran (1.04 g) and trifluoroacetic acid (94.22 mg) were refluxed in tetrahydrofuran (20 mL) for 5 h. After the reaction was completed, the reaction mixture was cooled to room temperature, added with water (20 mL), extracted with ethyl acetate (50 mL x 2), washed with saturated brine (10 mL x 3), dried over anhydrous sodium sulfate, filtered and dried by rotary evaporation, and the residue was purified by column chromatography (1000 mesh silica gel, petroleum ether/ethyl acetate = 100/1 to 5/1) to give compound 1E. LCMS(ESI)m/z: 278(M+1); 'HNMR(CHLOROFORM-d) 6 ppm 7.79-7.96(m, 1H), 5.25-5.32(m, 1H), 4.08-4.15(m, 1H), 3.66-3.79(m, 1H), 2.94-3.03(m, 2H), 2.65-2.78(m, 2H), 2.03-2.19(m, 6H), 1.73-1.79(m, 2H). Preparation of compound IG: Sodium (156 mg) was added to ethanol (9 mL) at room temperature, and after sodium completely disappeared, compound 1E (1.0 g) and compound IF (616 mg) were added to the resultant solution and refluxed for 5 h. After the reaction was completed, the reaction mixture was concentrated to dryness under reduced pressure, and the residue was purified by reversed-phase flash chromatography (Agilent, C18 reversed-phase column, 20-35 tm, 0.1% aqueous formic acid solution/acetonitrile) to give compound 1G. LCMS(ESI)m/z: 333(M+1); IHNMR(METHANOL-d4) 6 ppm 7.99(s, 1H), 5.27-5.46(m, 1H), 4.10-4.13(m, 1H), 3.97-4.09(m, 1H), 3.68-3.81(m, 1H), 2.94-3.15(m, 2H), 2.66-2.82(m, 2H), 1.92-2.17(m, 5H), 1.64(brs, 3H). Preparation of compound 1I: To a solution of compound IG (313.41 mg) and compound 1H (0.3 g) in dichloromethane (10 mL) was added diisopropylethylamine (349.92 mg) at 20 °C under nitrogen atmosphere. The reaction mixture was stirred at 20 °C for 5 h. After the reaction was completed, the reaction mixture was concentrated to dryness to give crude compound II for direct use in the next step. LCMS (ESI) m/z: 470 (M+1). Preparation of compound 1J: Compound 11(0.4238 g) was dissolved in a mixed solvent of methanol (10 mL) and water (10 mL) at room temperature, followed by addition of sodium hydroxide (360.96 mg). The resultant mixture was warmed to 70 °C and stirred for 10 min. After the reaction was completed, the reaction mixture was distilled to remove methanol, added with water (10 mL), extracted with ethyl acetate (50 mL x 2), washed with saturated brine (10 mL x 2), dried over anhydrous sodium sulfate, filtered and dried by rotary evaporation, and the residue was purified by reversed-phase flash chromatography (Agilent, C18 reversed-phase column, 20-35 [tm, 0.1% aqueous formic acid solution/acetonitrile) to give compound 1J. LCMS (ESI) m/z: 452 (M+1). Preparation of compound 1-1 hydrochloride and compound 1-2 hydrochloride: To a solution of compound 1J (0.23 g) in dichloromethane (6 mL) was added dropwise trifluoroacetic acid (4 mL) at room temperature, followed by stirring at room temperature for 1 h. After the reaction was completed, the reaction mixture was concentrated, and the residue was purified by reversed-phase flash chromatography (Agilent, C18 reversed-phase column, 20-35 tm, 0.1% aqueous formic acid/acetonitrile). The resultant product was separated by SFC (column: Daicel OD (250 mm x 30mm,10 tm); mobile phase: carbon dioxide as phase A, ethanol containing 0.1% aqueous ammonia as phase B; elution gradient: isocratic elution with 50% phase B, duration for each injection: 4 min), and the resultant two fractions were separately re-purified by reversed-phase flash chromatography (Agilent, C18 reversed-phase column, 20-35 [tm, 0.1% aqueous hydrochloric acid/acetonitrile) to give compound 1-1 hydrochloride and compound 1-2 hydrochloride. Determined using the SFC analytical method below, the retention times of the hydrochloride of compound 1-1 hydrochloride and compound 1-2 hydrochloride were 2.200 min and 2.663 min, respectively. SFC analytical method: Column: Daicel OD-350 x 4.6 mm I.D., 3 tm; mobile phase: carbon dioxide as phase A, ethanol containing 0.05% diethylamine as phase B; gradient elution: 5-40% phase B; flow rate: 3 mL/min; wavelength: 220 nm; column temperature: 35 °C; back pressure: 100 Bar. Compound 1-1 hydrochloride: LCMS(ESI)m/z: 368(M+1); 1 HNMR(DMSO-d6) 6 ppm 12.37-12.95(m, 1H), 9.96-10.23(m, 1H), 7.98(s, 1H), 4.76-4.83(m, 1H), 3.64-3.73(m, 1H), 3.41-3.42(m, 1H), 3.30-3.39(m, 2H), 3.16-3.25(m, 1H), 3.08(brs, 3H), 2.34-2.43(m, 1H), 2.11-2.27(m, 2H), 1.96-2.09(m, 2H), 1.74-1.95(m, 4H). Compound 1-2 hydrochloride: LCMS(ESI)m/z: 368(M+1); 1 HNMR(DMSO-d6) 6 ppm 12.44-13.09(m, 1H), 9.87-10.10(m, 1H), 7.98(s, 1H), 4.73-4.82(m, 1H), 3.69-3.80(m, 1H), 3.45-3.58(m, 1H), 3.28-3.42(m, 2H), 3.13-3.25(m, 1H), 3.04-3.13(m, 3H), 2.36-2.44(m, 1H), 2.12-2.25(m, 2H), 1.94-2.04(m, 2H), 1.74-1.93(m, 4H). Example 2. Compound 2-1 Hydrochloride and Compound 2-2 Hydrochloride 0

N-.C zb a1O H 2N00 N NH 2 2A \N S NH Cbz ,\S N 0 NH Cbz NNN N NH 2 o 2C 1G 2B 0 0

HN \ H HN \ S NH N N +NN

2-1 or 2-2 2-2 or 2-1

Preparation of compound 2B: The compound was prepared as described for compound 1I. LCMS (ESI) m/z: 564 (M+1). Preparation of compound 2C: The compound was prepared as described for compound 1J. LCMS (ESI) m/z: 546 (M+1). Preparation of compound 2-1 hydrochloride and compound 2-2 hydrochloride: To compound 3B (0.1 g) was added dropwise a 33% solution of hydrobromic acid in acetic acid (3 mL) at room temperature, followed by stirring at room temperature for 1 h. After the reaction was completed, the reaction mixture was concentrated, and the residue was purified by reversed-phase flash chromatography (Agilent, C18 reversed-phase column, 20-35 tm, 0.1% aqueous formic acid/acetonitrile) and then separated by SFC (column: Daicel AD (250 mm x 30 mm,10 tm); mobile phase: carbon dioxide as phase A, ethanol containing 0.1% aqueous ammonia as phase B; elution gradient: isocratic elution with 50% phase B, duration for each injection: 5.6 min). The resultant two fractions were separately re-purified by reversed-phase flash chromatography (Agilent, C18 reversed-phase column, 20-35 tm, 0.1% aqueous hydrochloric acid/acetonitrile) to give compound 2-1 hydrochloride and compound 2-2 hydrochloride.

Determined using the SFC analytical method below, the retention times of compound 2-1 and compound 2-2 were 0.932 min and 1.421 min, respectively. SFC analytical method: Column: Daicel OD-350 x 4.6 mm I.D., 3 tm; mobile phase: carbon dioxide as phase A, ethanol containing 0.05% diethylamine as phase B; gradient elution: isocratic elution with 40% phase B; flow rate: 3 mL/min; wavelength: 220 nm; column temperature: 35 °C; back pressure: 100 Bar. Compound 2-1 hydrochloride LCMS(ESI)m/z: 328(M+); 'HNMR(DMSO+D20) 6 ppm 7.93(s, 1H), 4.63-4.72(m, 1H), 3.41-3.53(m, 1H), 3.27-3.38(m, 1H), 3.09-3.20(m, 2H), 2.95-3.07(m, 2H), 2.39-2.50(m, 1H), 1.99-2.12(m, 3H), 1.87-1.99(m, 2H). Compound 2-2 hydrochloride LCMS(ESI)m/z: 328(M+); 'HNMR(DMSO+D20) 6 ppm 7.93(s, 1H), 4.63-4.72(m, 1H), 3.41-3.53(m, 1H), 3.27-3.38(m, 1H), 3.09-3.20(m, 2H), 2.95-3.07(m, 2H), 2.39-2.50(m, 1H), 1.99-2.12(m, 3H), 1.87-1.99(m, 2H). Example 3. Compound 3-1 Hydrochloride and Compound 3-2 Hydrochloride 0 0

HN\ S NH HN \ S NH N \ N N C) N N- b N N N

3-1or3-2 3-2 or3-1

Preparation of compound 3-1 hydrochloride: Compound 2-2 hydrochloride (50.00 mg), 37% aqueous formaldehyde (57 tL) and sodium cyanoborohydride (47.99 mg) were stirred in methanol 10 (mL) for 1 h. After the reaction was completed, the reaction mixture was concentrated, and the residue was purified by preparative high performance liquid chromatography (column: Phenomenex Synergi C18 150 x 25 x 10 [tm; mobile phase: 0.05% aqueous hydrochloric acid-acetonitrile; acetonitrile gradient: 6-26%, duration: 12 min) to give compound 3-1 hydrochloride. LCMS(ESI)m/z: 342(M+); 'HNMR(400MHz, DMSO+D20) 6 ppm 7.94(s, 1H), 4.55(brt, J=7.94Hz, 1H), 3.73-3.77(m, 1H), 3.24-3.37(m, 1H), 3.10-3.23(m, 2H), 3.03-3.08(m, 2H), 2.98(s, 3H), 2.67(brd, J=7.13Hz, 1H), 2.16(brd, J=7.75Hz, 1H), 1.91-2.08(m, 4H). Preparation of compound 3-2 hydrochloride: Compound 3-2 hydrochloride was prepared from compound 2-1 hydrochloride, as described for compound 3-1 hydrochloride. LCMS(ESI)m/z: 342(M+); HNMR(400MHz, DMSO+D20) 6 ppm 7.96(s, 1H), 4.55(brt, J=7.94Hz, 1H), 3.77-3.82(m, 1H), 3.31(m, 1H), 3.09-3.25(m, 2H), 3.04-3.09(m, 2H), 2.99(s, 3H), 2.63-2.71(m, 1H), 1.93-2.18(m, 5H). Determined using the SFC analytical method below, the retention times of compound 3-1 hydrochloride and compound 3-2 were 1.879 min and 1.788 min, respectively. SFC analytical method:

Column: Cellucoat OD-350 x 4.6 mm I.D., 3 tm; mobile phase: carbon dioxide as phase A, methanol containing 0.05% diethylamine as phase B; gradient elution: 5-40% phase B; flow rate: 3 mL/min; wavelength: 220 nm; column temperature: 35 °C; back pressure: 100 Bar. Example 4. Compound 4-1 Hydrochloride and Compound 4-2 Hydrochloride O H2 N H 2N

N NH 2 'Cbz 4A bz HN \%\O H N 00 N'~ 0 NH

SNH2

IG 4B 4C

0 0

HN\ H HN\ 3 NH N NHN N

4-1 or 4-2 4-2 or 4-1

Preparation of compound 4B: The compound was prepared as described for compound 1I. LCMS (ESI) m/z: 578. Preparation of compound 4C: To compound 4B (0.27 g) was added dropwise a 33% solution of hydrobromic acid in acetic acid (2 mL) at room temperature, followed by stirring at room temperature for 1 h. After the reaction was completed, the reaction mixture was concentrated to give compound 3C (bright yellow oil, 170 mg) for direct use in the next step. LCMS (ESI) m/z: 360. Preparation of compound 4-1 hydrochloride and compound 4-2 hydrochloride: Compound 4C (170 mg) and sodium hydroxide (189 mg) were stirred in a mixed solution of methanol (3 mL) and water (1 mL) at 70 °C for 0.5 h. After the reaction was completed, the reaction mixture was concentrated, and the residue was purified by preparative high performance liquid chromatography (column: Phenomenex Synergi C18 150 x 25 x 10 tm; mobile phase: 0.05% aqueous hydrochloric acid-acetonitrile; acetonitrile gradient: 8-28%, duration: 11min) and then separated by SFC (column: Daicel IG (250 mm x 50mm,10 tm); mobile phase: carbon dioxide as phase A; methanol solution containing 0.1% aqueous ammonia as phase B; elution gradient: isocratic elution with 55% phase B; duration for each injection: 4.0 min) to give two fractions. Fraction 1 was re-purified by preparative high performance liquid chromatography (column: Phenomenex Synergi C18 150 x 25 x 10 tm; mobile phase: 0.05% aqueous hydrochloric acid-acetonitrile; acetonitrile gradient: 7-27%, duration: 11 min) to give compound 4-1 hydrochloride (100% ee), and fraction 2 was used as compound 4-2 (99.220% ee) without purification. Compound 4-1 hydrochloride: LCMS(ESI)m/z: 342(M+1); 'HNMR(400MHz, DMSO-d6) 6 ppm 12.77(brs, 1H), 10.07(brs, 1H), 9.09(brs, 1H), 7.97(s, 1H), 3.40(brd, J=5.OlHz, 2H), 3.19(brd, J=2.8lHz, 2H), 3.08(brd, J=5.26Hz, 2H), 2.33-2.39(m, 1H), 2.18-2.25(m, 1H), 2.03-2.11(m, 1H), 1.98(brs, 2H), 1.83-1.91(m, 1H), 1.78(s, 3H). Compound 4-2: LCMS(ESI)m/z: 342(M+1); 1 HNMR(400MHz, DMSO-d6) 6 ppm 7.93(s, 1H), 3.24(brd, J=8.07Hz, 1H), 3.11-3.18(m, 3H), 3.05-3.09(m, 2H), 2.64-2.73(m, 1H), 2.35-2.43(m, 1H), 1.92-2.04(m, 4H), 1.78(brd, J=7.09Hz, 1H), 1.65(s, 3H).

Determined using the SFC analytical method below, the retention times of compound 4-1 hydrochloride and compound 4-2 were 1.777 min and 2.687 min, respectively. SFC analytical method: Column: Daicel IG-350 x 4.6 mm I.D., 3 tm; mobile phase: carbon dioxide as phase A, methanol containing 0.05% diethylamine as phase B; gradient elution: isocratic elution with 40% phase B; flow rate: 3 mL/min; wavelength: 220 nm; column temperature: 35 °C; back pressure: 100 Bar. Example 5. Compound 5-1 Hydrochloride and Compound 5-2 Hydrochloride o O HN S NH HN S NH N-. \ N-. \'~ N N NN N

5-1 or 5-2 5-2 or 5-1

Preparation of compound 5-1 hydrochloride and compound 5-2 hydrochloride: The compounds were prepared from compound 4-1 hydrochloride and compound 4-2 hydrochloride, respectively, as described in Example 3. 1 Compound 5-1 hydrochloride: LCMS(ESI)m/z: 356(M+1); HNMR(400MHz, METHANOL-d4) 6 ppm 8.08(s, 1H), 3.89-4.05(m, 1H), 3.45-3.55(m, 1H), 3.34-3.43(m, 1H), 3.11-3.28(m, 6H), 2.51-2.64(m, 1H), 2.38-2.49(m, 1H), 2.32(brd, J=8.8OHz, 1H), 2.05-2.22(m, 3H), 1.85(s, 3H). 1 Compound 5-2 hydrochloride: LCMS(ESI)m/z: 356(M+1); HNMR(400MHz, METHANOL-d4) 6 ppm 8.17(s, 1H), 3.89-4.02(m, 1H), 3.45-3.54(m, 1H), 3.35-3.42(m, 1H), 3.10-3.27(m, 6H), 2.51-2.64(m, 1H), 2.38-2.49(m, 1H), 2.32(brd, J=8.8OHz, 1H), 2.08-2.21(m, 3H), 1.85(s, 3H). Determined using the SFC analytical method below, the retention times of compound 5-1 hydrochloride and compound 5-2 hydrochloride were 2.148 min and 1.907 min, respectively. SFC analytical method: Column: Daicel OD-350 x 4.6 mm I.D., 3 tm; mobile phase: carbon dioxide as phase A, methanol containing 0.05% diethylamine as phase B; gradient elution: 5-40% phase B; flow rate: 3 mL/min; wavelength: 220 nm; column temperature: 35 °C; back pressure: 100 Bar. Example 6. Compound 6-1 Hydrochloride and Compound 6-2 Hydrochloride o O

HN S NH N S NH N N N N N

6-1 or 6-2 6-2 or 6-1

Preparation of compound 6-1 hydrochloride: Example 2-1(95 mg), acetone (107 tL) and sodium cyanoborohydride (91 mg) were stirred in methanol (10 mL) at room temperature for 1 h. After the reaction was completed, the reaction mixture was concentrated, and the residue was purified by preparative high performance liquid chromatography (column: Phenomenex synergy C18 150 x 25 x 10 [tm; mobile phase: 0.05% aqueous hydrochloric acid-acetonitrile; acetonitrile gradient: 11-31%, duration: 11 min) to give compound 6-1 hydrochloride. LCMS(ESI)m/z: 370(M+1); IHNMR(400MHz, DMSO-d6) 6 ppm 12.75-13.15(m, 1H), 9.63(brs, 1H), 7.97(s, 1H), 4.72(brd, J=7.70Hz, 1H), 3.82(brdd, J=6.48, 4.03Hz, 1H), 3.67-3.73(m, 1H), 3.33-3.47(m, 1H), 3.19-3.33(m, 1H), 3.04-3.15(m, 3H), 2.56-2.70(m, 1H), 1.92-2.18(m, 5H), 1.23-1.35(m, 6H). Preparation of compound 6-2 hydrochloride: Compound 6-2 hydrochloride was prepared from compound 2-2 hydrochloride, as described for compound 6-1 hydrochloride. LCMS(ESI)m/z: 370(M+1); 1 HNMR(400MHz, DMSO-d6) 6 ppm 12.59-13.14(m, 1H), 9.63(brs, 1H), 7.97(s, 1H), 4.63-4.77(m, 1H), 3.82(brdd, J=6.48, 3.79Hz, 1H), 3.71(brd, J=6.36Hz, 1H), 3.34-3.49(m, 1H), 3.19-3.33(m, 1H), 3.03-3.14(m, 3H), 2.53-2.66(m, 1H), 2.18-2.36(m, 1H), 1.90-2.15(m, 5H), 1.32(d, J=6.60Hz, 3H), 1.26(d, J=6.60Hz, 3H). Determined using the SFC analytical method below, the retention times of compound 6-1 hydrochloride and compound 6-2 hydrochloride were 1.838 min and 1.992 min, respectively. SFC analytical method: Column: Daicel OD-350 x 4.6 mm I.D., 3 tm; mobile phase: carbon dioxide as phase A, methanol containing 0.05% diethylamine as phase B; gradient elution: 5-40% phase B; flow rate: 3 mL/min; wavelength: 220 nm; column temperature: 35 °C; back pressure: 100 Bar. Example 7. Compound 7-1 Formate and Compound 7-2 Formate

O N 07A ONH N H2 O1HN\ SH SN \DONN N, O N

NH 2 7B IG

0 0 HN\ S NH HN\ S NH SN N H N

7-1 or 7-2 7-2 or 7-1

Preparation of compound 7B: To a solution of compound 7A (91.72 mg) and diisopropylethylamine (106.99 mg) in dichloromethane (2 mL) was added 2,2-dicarbonylimidazole (44.74 mg) at room temperature, followed by stirring at room temperature for 2 h. The reaction mixture was distilled to remove the solvent, and the residue was added with compound 6A (100 mg) and N,N-dimethylformamide (2 mL), followed by stirring at 100 °C for 48 h. After the reaction was completed, the reaction mixture was concentrated to dryness, and the residue was purified by reversed-phase flash chromatography (Agilent, C18 reversed-phase column, 20-35 tm, 0.1% aqueous formic acid solution/acetonitrile) to give compound 7B formate. LCMS (ESI) m/z: 440. Preparation of compound 7-1 formate and compound 7-2 formate: The compounds were prepared as described in Example 1.

Compound 7-1 formate: LCMS(ESI)m/z: 356(M+1); 'HNMR(METHANOL-d4) 6 ppm 8.09(s, 1H), 4.20-4.23(m, 1H), 3.67-3.76(m, 1H), 3.29(brd, J=4.0Hz, 2H), 3.18(brd, J=5.7Hz, 2H), 2.97(s, 3H), 2.32-2.41(m, 1H), 2.10-2.20(m, 2H), 1.99(s, 5H), 1.69-1.85(m, 1H). Compound 7-2 formate: LCMS(ESI)m/z: 356(M+1); 'HNMR(METHANOL-d4) 6 ppm 8.06(s, 1H), 4.19-4.27(m, 1H), 3.67-3.76(m, 1H), 3.29(brd, J=4.0Hz, 2H), 3.18(brd, J=5.7Hz, 2H), 2.97(s, 3H), 2.32-2.41(m, 1H), 2.10-2.20(m, 2H), 1.99(s, 5H), 1.69-1.85(m, 1H). Determined using the SFC analytical method below, the retention times of compound 7-1 formate and compound 7-2 formate were 0.943 min and 1.326 min, respectively. SFC analytical method: Column: Chiralpak AD-350 x 4.6 mm I.D., 3 tm; mobile phase: carbon dioxide as phase A, ethanol containing 0.05% diethylamine as phase B; gradient elution: isocratic elution with 40% phase B; flow rate: 3 mL/min; wavelength: 220 nm; column temperature: 35 °C; back pressure: 100 Bar. Example 8. Compound 8-1 Hydrochloride and Compound 8-2 Hydrochloride

ON CI S H2N 0 H2 Cbz' ,\ S 8A NH O N'Cbz 0 NH 2

IG 8B

00 0

NHN N HN S NH NH N N N:Z \1 N'Cbz I- : 5 N-- NH +N NH

8-1 or 8-2 8-2 or 8-1 8C

Preparation of compound 8B: The compound was prepared as described for compound 1I. LCMS (ESI) m/z: 578. Preparation of compound 8C: The compound was prepared as described for compound 1J. LCMS (ESI) m/z: 560. Preparation of compound 8-1 hydrochloride and compound 8-2 hydrochloride: Prepared as described in Example 2. Compound 8-1 hydrochloride: LCMS(ESI)m/z: 342(M+1); 1 HNMR(400MHz, DMSO-d6) 6 ppm 12.47(brs, 1H), 9.12-9.39(m, 2H), 7.94(s, 1H), 3.47(brd, J=11.OOHz, 1H), 3.15-3.30(m, 3H), 3.02-3.12(m, 4H), 2.87-2.96(m, 1H), 2.14(brd, J=10.15Hz, 1H), 1.93-2.02(m, 2H), 1.75-1.88(m, 2H), 1.61-1.72(m, 1H). Compound 8-2 hydrochloride: LCMS(ESI)m/z: 342(M+1); 1 HNMR(400MHz, DMSO-d6) 6 ppm 12.45(brs, 1H), 9.05-9.47(m, 2H), 7.95(s, 1H), 3.47(brd, J=10.88Hz, 1H), 3.15-3.31(m, 3H), 3.01-3.13(m, 4H), 2.85-2.97(m, 1H), 2.14(brd, J=9.78Hz, 1H), 1.92-2.02(m, 2H), 1.75-1.89(m, 2H), 1.63-1.72(m, 1H). Determined using the SFC analytical method below, the retention times of compound 8-1 hydrochloride and compound 8-2 hydrochloride were 0.902 min and 1.802 min, respectively. SFC analytical method:

Column: Chiralpak AD-350 x 4.6 mm I.D., 3 tm; mobile phase: carbon dioxide as phase A, ethanol containing 0.05% diethylamine as phase B; gradient elution: isocratic elution with 40% phase B; flow rate: 3 mL/min; wavelength: 220 nm; column temperature: 35 °C; back pressure: 100 Bar. Example 9. Compound 9-1 Hydrochloride and Compound 9-2 Hydrochloride 0 0

HN\ S NH HN\ S NH N \ - N NI NN A N

' 9-1 or 9-2 9-2 or 9-1

Preparation of compound 9-1 hydrochloride and compound 9-2 hydrochloride: The compounds were prepared from compound 8-1 hydrochloride and compound 8-2 hydrochloride, respectively, as described in Example 3-1. Compound 9-1 hydrochloride: LCMS(ESI)m/z: 356(M+1); 'HNMR(400MHz, DMSO-d6) 6 ppm 10.97(brs, 1H), 7.95(s, 1H), 3.57-3.64(m, 1H), 3.29-3.43(m, 3H), 3.04-3.12(m, 4H), 2.90-2.99(m, 1H), 2.80(d, J=4.65Hz, 3H), 2.17(brd, J=13.45Hz, 1H), 1.87-1.99(m, 3H), 1.36-1.69(m, 1H). Compound 9-2 hydrochloride: LCMS(ESI)m/z: 356(M+1); 1 HNMR(400MHz, DMSO-d6) 6 ppm 10.97(brs, 1H), 7.95(s, 1H), 3.61(brd, J=7.34Hz, 1H), 3.31-3.42(m, 3H), 3.03-3.10(m, 4H), 2.90-2.99(m, 1H), 2.80(d, J=4.77Hz, 3H), 2.17(brd, J=13.45Hz, 1H), 1.90-2.00(m, 4H), 1.43-1.64(m, 1H). Determined using the SFC analytical method below, the retention times of compound 9-1 hydrochloride and compound 9-2 hydrochloride were 2.183 min and 1.681 min, respectively. SFC analytical method: Column: Chiralpak IC-350 x 4.6 mm I.D., 3 tm; mobile phase: carbon dioxide as phase A, ethanol containing 0.05% diethylamine as phase B; gradient elution: isocratic elution with 40% phase B; flow rate: 3 mL/min; wavelength: 220 nm; column temperature: 35 °C; back pressure: 100 Bar. Example 10. Compound 10 Hydrochloride

0 H 2N O ~ Cbz-NDN4H2 CICZN S 0 N NH 2 IOA \ S N NH NJ 0 1 NH 2 NCb N'Cbz

O O

S NH HN S NH N. N N NCbz 10C 10

Preparation of compound 1OB: The compound was prepared as described for compound 1I. LCMS (ESI) m/z: 578. Preparation of compound 1OC: The compound was prepared as described for compound 1J. LCMS (ESI) m/z: 560. Preparation of compound 10 hydrochloride: The compound was prepared as described for compound 2-1 hydrochloride. LCMS(ESI)m/z: 342(M+1); IHNMR(400MHz, DMSO-d6) 6 ppm 12.12 - 12.63 (m, 1 H), 9.01 (br d, J=8.93 Hz, 1 H), 8.65 (br d, J=9.78 Hz, 1 H), 7.94 (s, 1 H), 3.40 (br d, J=6.72 Hz, 1 H), 2.91 - 3.08 (m, 7 H), 1.91 - 2.17 (m, 7 H). Example 11. Compound 11 Hydrochloride 0 0

HN S NH HN S NH N.\ N NNA NNN NN N NH N

10 11 Preparation of compound 11 hydrochloride: The compound was prepared from compound 10 hydrochloride, as described in Example 3-1. LCMS(ESI)m/z: 1 356(M+1); HNMR(400MHz, DMSO-d6) 6 ppm 12.37(brs, 1H), 10.41(brs, 1H), 7.84, -8.07(m, 1H), 3.47-3.56(m, 1H), 3.49(brd, J=11.25Hz, 1H), 3.19-3.36(m, 1H), 2.96-3.17(m, 6H), 2.82-2.95(m, 1H), 2.76(d, J=4.65Hz, 3H), 2.05-2.32(m, 4H), 1.91-2.04(m, 2H), 1.06(t, J=7.03Hz, 1H). Example 12. Compound 12 Hydrochloride

Cbz-NQN 0 H 2 N CI N S1 0 N NH 2 12A O N _Cbz NH '\ S 0 \INH 2 5t NCb IG 12B

O O S NH HN S NH NNH NN N NHI 'Cbz

12C 12

Preparation of compound 12 hydrochloride: The compound was prepared as described for compound 2-1 hydrochloride. LCMS (ESI) m/z: 356 (M+). IHNMR (400 MHz, DMSO-d) 6 ppm 12.14 (brs, 1H), 8.78 (brs, 2H), 7.86-8.03 (m, 1H), 2.99-3.26 (m, 8H), 2.52-2.56 (m, 2H), 1.73-2.06 (m, 4H), 1.27-1.42 (m, 3H).

Example 13. Compound 13 Hydrochloride 0 0 HN S NH HN S NH

N N NH

12 13

Preparation of compound 13 hydrochloride: The compound was prepared from compound 12 hydrochloride, as described for compound 3-1 hydrochloride. LCMS(ESI)m/z: 370(M+1); 1 HNMR(400MHz, DMSO-d6) 6 ppm 11.94-12.36(m, 1H), 10.16-10.60(m, 1H), 7.82-8.04(m, 1H), 3.32(brd, J=11.98Hz, 2H), 2.98-3.11(m, 5H), 2.61-2.80(m, 4H), 2.09-2.32(m, 2H), 1.81-2.05(m, 4H), 1.44(s, 3H). Example 14. Compound 14-1 Hydrochloride and Compound 14-2 Hydrochloride

NH 2 O S NH 2 O 14A NH 2 O , CI NH ,-N Oo N

IG 14B

0 0 HN\ S NH HN S NH N N5

14-1 or 14-2 14-2 or 14-1 Example 14 was prepared as described in Example 1. Compound 14-1 hydrochloride: LCMS(ESI)m/z: 354(M+1); 1 HNMR(400MHz, DMSO-d6) 6 ppm2.78(brs, 1H), 11.01(brs, 1H), 7.98(s, 1H), 4.88(s, 1H), 3.92(brd, J=2.6Hz, 1H), 3.50(brd, J=2.4Hz, 1H), 3.42-3.29(m, 3H), 3.20-3.03(m, 4H), 2.15(brd, J=2.8Hz, 1H), 2.05-1.87(m, 3H), 1.83-1.71(m, 2H). Compound 14-2 hydrochloride: LCMS(ESI)m/z: 354(M+1); 1 HNMR(400MHz, DMSO-d6) 6 ppm 12.78(brs, 1H), 11.01(brs, 1H), 7.98(s, 1H), 4.88(s, 1H), 3.92(brd, J=2.6Hz, 1H), 3.50(brd, J=2.4Hz, 1H), 3.42-3.29(m, 3H), 3.20-3.03(m, 4H), 2.15(brd, J=2.8Hz, 1H), 2.05-1.87(m, 3H), 1.83-1.71(m, 2H). Determined using the SFC analytical method below, the retention times of compound 14-1 hydrochloride and compound 14-2 hydrochloride were 1.453 min and 2.828 min, respectively. SFC analytical method: Column: Chiralpak AD-350 x 4.6 mm I.D., 3 tm; mobile phase: carbon dioxide as phase A, methanol containing 0.05% diethylamine as phase B; gradient elution: isocratic elution with 40% phase B; flow rate: 3 mL/min; wavelength: 220 nm; column temperature: 35 °C; back pressure: 100 Bar. Experimental Example 1. Detection of Inhibitory Effect of Compounds Against Activity of Cdc7/DBF4 Materials: Cdc7/DBF4 kinase detection kit purchased from Promega; and

Nivo multi-marker analyzer (PerkinElmer). Method: An enzyme, a substrate, adenosine triphosphate and an inhibitor were diluted with the kinase buffer in the kit. A test compound was serially 5-fold diluted to an 8th concentration, i.e., from 10 tM to 0.13 nM, with the DMSO concentration being 5%, and the duplicate well experiment was set up. To a microplate were added 1 pL of inhibitors of various concentration gradients, 2 pL of CDC7/DBF4 kinase (6.25 ng), 2 pL of a mixture of substrate and ATP (10 tM adenosine triphosphate, 0.2 tg/pL substrate), and the final concentration gradient of the compound was diluted from 2 tM to 0.025 nM. The reaction system was left reacting at 25 °C for 60 min. After the reaction was completed, 5 tL of ADP-Glo reagent was added to each well, and the reaction was continued at 25 °C for 40 min. After the reaction was completed, 10 tL of the kinase detection reagent was added to each well. After 30 min of reaction at 25 °C, the chemiluminescence was read using the multi-marker analyzer with an integration time of 0.5 s. Data analysis: The original data were converted to inhibition using the equation (Sample - Min)/(Max - Min) x 100%, and the IC50 value was then curve fitted using four parameters (obtained from the "log(inhibitor) vs. response-Variableslope" model in GraphPadPrism). Table 1 provides the enzymatic inhibitory activity of the compounds disclosed herein against Cdc7/DBF4. Results: See Table 1. Table 1 Cdc7/DBF4 IC5 0 Cdc7/DBF4 IC50 Test compounds (nmol) Testcompounds (nmol) Compound11 3.02 Compound 7-1 formate 3.07 hydrochloride Compound 1-2 1.29 Compound 7-2 format 2.7 hydrochloride Compound 2-1 3.73 Compound 8-1 4.27 hydrochloride hydrochloride Compound2-2 1.52 Compound 8-2 1.17 hydrochloride hydrochloride Compound 3-1 1.19 Compound 9-1 4.85 hydrochloride hydrochloride Compound 3-2 0.42 Compound 9-2 1.09 hydrochloride hydrochloride Compound 4-1 4.56 Compound 10 1.69 hydrochloride hydrochloride Compound 4-2 6.73 Compound 11 4.85 hydrochloride hydrochloride Compound 5-1 2.04 Compound 12 1.38 hydrochloride hydrochloride Compound5-2 2.35 Compound 13 5.66 hydrochloride hydrochloride Compound 6-1 1.7 Compound 14-1 1.21 hydrochloride hydrochloride Compound 6-2 0.99 Compound 14-2 1.43 hydrochloride ____________hydrochloride ___________

Experimental Example 2. Detection of Inhibitory Effect of Compounds Against Activity of Colo205 Cells

Materials: 1640 medium; fetal bovine serum; penicillin/streptomycin antibiotics purchased from Wisent; CellTiter-Glo (chemiluminescence detection reagent for cell viability) reagent purchased from Promega; COL0205 cell line purchased from Wuhan Procell Life Science&Technology Co., Ltd; and Nivo multi-marker analyzer (PerkinElmer). Method: COL0205 cells were plated on to white 96-well plates by adding 80 pL of cell suspension (containing 3000 COL0205 cells) to each well. The cell plate was incubated in a CO 2 incubator overnight. A test compound was serially 3-fold diluted to an 8th concentration, i.e., from 2 mM to 920 nM, and the duplicate well experiment was set up. 78 pL of medium was added to an intermediate plate, 2 pL of the serially diluted compound was transferred to corresponding wells of the intermediate plate, and after mixing, the mixture was transferred to the cell plate at 20 pL per well. The concentration of the compound transferred to the cell plate ranged from 10 pM to 4.57 nM. The cell plate was incubated in a CO 2 incubator for 3 days. Another cell plate was provided for reading signal values on the day of compound addition and these values were used as Max values (the Max in the equation below) in data analysis. The chemiluminescence detection reagent for cell viability was added to this cell plate at 25 pL per well and the luminescence signals were stabilized by incubation at room temperature for 10 min. Readings were taken using the multi-marker analyzer. The chemiluminescence detection reagent for cell viability was added to the cell plate at 25 pL per well and the luminescence signals were stabilized by incubation at room temperature for 10 min. Readings were taken using the multi-marker analyzer. Data analysis: The original data were converted to inhibition using the equation (Sample - Min)/(Max - Min) x 100%, and the IC50 value was then curve fitted using four parameters (obtained from the "log(inhibitor) vs. response-Variableslope" model in GraphPadPrism). Table 2 provides the inhibitory activity of the compounds disclosed herein against COL0205 cell proliferation. Results: See Table 2. Table 2

Test compounds Colo205 IC50 (nmol)

Compound 1-1 hydrochloride 13.8

Compound 1-2 hydrochloride 53.62

Compound 14-1 hydrochloride 50.09

Experimental Example 3. Single-Dose Pharmacokinetic Study in Mice

Objective: To evaluate the pharmacokinetic behavior by using male CD-i mice as test animals and determining the drug concentrations of the compounds in the plasma after single-dose administration. Method:

Healthy adult male CD-i mice were selected for intragastric administration. A compound was mixed with an appropriate amount of 5% DMSO/95% (10% hydroxypropyl-p-cyclodextrin), vortexed and sonicated to prepare a 1 mg/mL clear solution for later use. After the mice were administered intravenously at 2 mg/kg and orally at 10 mg/kg, whole blood was collected at certain time points, and plasma was separated. After pretreatment of the samples, the drug concentration was measured by LC-MS/MS, and pharmacokinetic parameters were calculated using Phoenix WinNonlin software. Results: See Table 3. Table 3. Pharmacokinetic results in mice

Pharmacokinetic results Compound 1-1 hydrochloride (IV: 2 mg/kg; PO: 10 mg/kg)

Clearance (mL/min/kg) 35.2

Apparent volume of distribution (L/kg) 4.13

AUCo-iast (intravenous injection, nM. h) 2454

AUCo-iast (oral, nM. h) 9080

Half-life (h) 4.03

Maximum concentration (nM) 2230

Bioavailability (%) 71.4

Experimental Example 4. In Vivo Pharmacodynamic Study of Compounds in Subcutaneous Xenograft Tumor Nude Mouse Model of Human Colorectal Cancer Cell SW620 Cell culturing: Human colorectal cancer cell SW620 cells of the 7th passage were cultured in an L-15 medium containing 10% fetal bovine serum, 100 U/mL penicillin and 100 pg/mL streptomycin through in vitro monolayer culture in an incubator at 37 °C/0% CO 2 for 4 passages with conventional medium replacement. At a cell saturation of

8 0- 9 0 %, the cells were digested with pancreatin-EDTA, counted and resuspended in PBS at a density of 5 x 106 cells/100 pL. Tumor cell inoculation and grouping: Cell inoculation: Each mouse was inoculated on the right cervical dorsum with 0.1 mL of the cell suspension of 5 x 106 SW620 cells in PBS. At an average tumor volume up to about 134 mm3, the mice were randomly grouped and administrated. Table 4. Animal grouping and administration regimen

Groups n Compounds Dosages (mg/kg) and Volumes Routes frequencies (pL/g) 1 6 Vehicle BID 10 p.o. 2 6 TAK-931 40 x BID 10 p.o. 3 6 Example 1-1 2.5 x BID 10 p.o. 4 6 Example 1-1 5 x BID 10 p.o. 5 6 Example 1-1 7.5 x BID 10 p.o. Note: Vehicle refers to a vehicle control group.

Tumor measurement and experimental indices: Tumor diameters were measured twice weekly using a vernier caliper. The tumor volume was calculated using 2 the following formula: V = 0.5 a x b2, where a and b represent the long diameter and short diameter of the tumor, respectively. The anti-tumor therapeutic effect of the compound was evaluated by TGI (%) or relative tumor proliferation rate T/C (%). Relative tumor proliferation rate T/C (%) = TRTV / CRTV x 100% (TRTV: RTV of treatment

group; CRTV: RTV of negative control group). Relative tumor volumes (RTVs) were calculated from the tumor measurement results using the following formula: RTV = Vt / VO, where VO is the average tumor volume measured at the time of grouping (i.e., DO), Vt is the average tumor volume measured at a certain measurement, and TRTV and CRTV are taken from the data measured on the same day. TGI (%), reflects the tumor growth inhibition rate. TGI (%) = [(1 - (average tumor volume at the end of

administration in a treatment group - average tumor volume at the start of administration of the treatment group)) / (average tumor volume at the end of treatment of the vehicle control group - average tumor volume at the start of treatment of the vehicle control group)] x 100%. After day 22 of the grouping, mice were euthanized, plasma and tumors were sampled and tumors were weighed and photographed. Results: Table 5, FIG. 1 and FIG. 2. Table 5. Antitumor effect of the compounds on the xenograft tumor model of the human colorectal cancer cell SW620 Tumor Tmrvlm volume Tumorlume RTV TGI(%) T/C(%) Groups (mm )

(Day1) (Day21) (Day21) (Day21) (Day21)

Vehicle, p.o, BID 134±14a 1112±122 - -

TAK-931, 134±13 176±27 1.36 96 16 40 mg/kg, BID13±3162 Compound 1-1 hydrochloride, 134±13 237±27 1.88 89 22 2.5 mg/kg, BID Compound 1-1 hydrochloride, 134±15 133±32 0.93 100 11 5 mg/kg, BID Compound 1-1 hydrochloride, 134±14 53±16 0.31 108 4 7.5 mg/kg, BID

Claims (15)

1. CLAIMS 1. A compound of formula (I), an isomer thereof or a pharmaceutically acceptable salt thereof,

   HNq S NH NN- \ N RN L R4 (I)

   wherein, the carbon atom with"* can be a chiral carbon atom present in a form of a single (R) or (S) enantiomer or in a form rich in one enantiomer; L is selected from the group consisting of -CH2 -CH 2-CH 2-, -CH 2-0-CH 2 -, -CH 2-S-CH 2 -, -CH 2-NH-CH 2 -, -NH-CH 2 -CH 2-, -S-CH 2 -CH2 - and -O-CH2 -CH 2-; R 1 is selected from the group consisting of H, halogen, CN, C 1 _6 alkyl, C3 6_ cycloalkyl, phenyl, and 5-6 membered heteroaryl, wherein the C 1 _6 alkyl, C3 _6 cycloalkyl, phenyl, and 5-6 membered heteroaryl are each independently optionally substituted with 1, 2 or 3 Ra, the 5-6 membered heteroaryl containing 1, 2 or 3 heteroatoms or heteroatom groups each independently selected from the group consisting of 0, S, N and NH; R2 is selected from Rb, R 3 is selected from NH 2 , and R 4 is selected from H; alternatively, R 2 is selected from R, and R3 and R 4 are joined to form a ring A group optionally substituted with 1, 2 or 3 R, wherein the ring A group is selected from the group consisting of C6_ 14 aryl, 5-14 membered heteroaryl, 5-12 membered heterocycloalkenyl and 4-14 membered heterocycloalkyl each independently containing 1, 2 or 3 heteroatoms or heteroatom groups independently selected from the group consisting of0, S, N and NRd;

   Ra is each independently selected from the group consisting of F, Cl, Br, I, OH, CN, NH 2 , -CH 3 and - 1; Rb is selected from the group consisting of H and C 1 6_ alkyl, the C 1 _6 alkyl being optionally substituted with 1, 2 or 3 Rbb; Re is selected from the group consisting of H, F, Cl, Br, I and C 1 _3 alkyl; Rd is selected from the group consisting of H and C 1 _4 alkyl; Rbb is selected from the group consisting of -OCH 3 , -OCH2 CH3 , -O-CH(CH 3) 2, cyclopropyl, cyclopentyl,

   phenyl, pyrazolyl, pyridyl, NH 2 , -NHCH 3 and -N(CH 3) 2; Re is selected from the group consisting of F, Cl, Br, I, OH, CN, COOH, NH2 , -NHCH 3, -N(CH 3)2 , -CH 3 ,

   -CH 2CH 3, -CF 3 , -OCH 3, -OCH 2CH 3 , -O-CH(CH 3)2 , -C(=0)OCH 3 , -C(=0)CH 3 and -C(=0)CH2CH 3 .
2. 2. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 1, wherein R 1 is selected from the group consisting of H, F, Cl, Br, I, CN, C1 _3 alkyl, C 35 cycloalkyl, phenyl, and 6 membered heteroaryl, wherein the C1 _3 alkyl, C 3 _ 5 cycloalkyl, phenyl, and 6 membered heteroaryl are each independently optionally substituted with 1, 2 or 3 Ra, and the 6 membered heteroaryl contains 1, 2 or 3 heteroatoms selected from N; preferably, R 1 is selected from the group consisting of H, F, Cl, Br, I, CN, -CH 3 ,

   -CH 2CH 3, cyclopropyl, phenyl and pyridyl, wherein the -CH 3 , -CH 2CH 3 , cyclopropyl, phenyl and pyridyl are each independently optionally substituted with 1, 2 or 3 Ra; further preferably, R1 is selected from the group consisting of H, F, Cl, Br,1, CN, CH 3 , CH 2CH 3, CF3 , cyclopropyl, phenyl and pyridyl; further more preferably, R 1 is selected from H.
3. 3. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 1, wherein Rb is selected from the group consisting of H and C14 alkyl, the C14 alkyl being optionally substituted with 1, 2 or 3 Rbb; preferably, Rb is selected from the group consisting of H, methyl, ethyl, isopropyl, n-propyl, n-butyl and isobutyl; further preferably, Rb is selected from C1_3 alkyl; further more preferably, Rb is selected from isopropyl.
4. 4. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 1, wherein Re is selected from the group consisting of H, F and C1_3 alkyl; preferably, Re is selected from the group consisting of H, methyl, ethyl and F; further preferably, Re is selected from the group consisting of H and methyl.
5. 5. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 1, wherein Rd is selected from the group consisting of H, methyl, ethyl, n-propyl, isopropyl, and n-butyl; preferably, Rd is selected from the group consisting of H and C1_3 alkyl; further preferably, Rd is selected from the group consisting of H, methyl and isopropyl.
6. 6. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 1, wherein the ring A group is selected from the group consisting of C6 1 o aryl, 5-9 membered heteroaryl, 5-7 membered heterocycloakenyl and 4-10 membered heterocycloalkyl optionally substituted with 1, 2 or 3 Re and each independently containing 1, 2 or 3 heteroatoms or heteroatom groups independently selected from the group consisting of 0, S, N and NR; preferably, the ring A group is selected from 5-9 membered heterocycloakyl optionally substituted with 1, 2 or 3 Re and containing 1, 2 or 3 heteroatoms or heteroatom groups independently selected from the group consisting of 0, S, N and NR; further preferably, the ring A group is selected from 5-9 membered heterocycloalkyl containing 1 heteroatom or heteroatom group selected from the group consisting of N and NRd.
7. 7. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 6, wherein the ring A group is selected from the group consisting of pyrrolidinyl, piperidinyl, morpholinyl, 1-azabicyclo[2.2.2]octanyl, 1-azabicyclo[2.2.1]heptanyl, 1-azabicyclo[3.2.2]nonanyl and azepanyl optionally substituted with 1, 2 or 3 Re, and contains 1 heteroatom or heteroatom group selected from the group N consisting of N and NRd; preferably, the ring A group is selected from the group consisting of

   Rd Rd I N jN <N < N - -- and
8. 8. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 1,

   R2 s*Rc wherein the structural unit R4 is selected from R4 , further from the group consisting of

   Rd Rd ,RC RR Rd Rd RRd R ad NN N Nd RcK

   XD I --I I and
9. 9. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 8,

   R2 *Re, R3 R3 wherein the structural unit R4 is selected from R4 , further from the group consisting of H H N H N N j<N NmN N

   N - H

   N N NH N

   R2 R3 NH 2 alternatively, the structural unit R4 is selected from, further from Rb , further more from

   NH 2
10. 10. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 1, wherein, the carbon atom with "*" can be a chiral carbon atom present in a form of a single (R) or (S) enantiomer or in a form rich in one enantiomer; L is selected from -CH2 -CH 2-CH 2 -; R 1 is selected from H; R2 is selected from Rb, R 3 is selected from NH 2 , and R 4 is selected from H; alternatively, R 2 is selected from R, and R3 and R4 are joined to form a ring A group selected from 4-14 membered heterocycloakyl each independently containing 1, 2 or 3 heteroatoms or heteroatom groups independently selected from the group consisting of N and NRd;

    Rb is selected from C1 _6 alkyl; Re is selected from the group consisting of H and C1 _3 alkyl; Rd is selected from the group consisting of H and C1 _4 alkyl.
11. 11. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 1, wherein the compound is selected from the group consisting of a compound of formula (1-1) and a compound of formula (1-2), an isomer thereof and a pharmaceutically acceptable salt thereof,

    '27

    HN\ S HN S NH~ \c NH Rb -.. I N N A N * NH2

    (I-i) Q-2)

    wherein the carbon atom with"*" can be achiral carbon atom present in aform of asingle (R) or () enantiomer or in aform rich in one enantiomer; Ris asdefined in claim 1; the ring Agroup is asdefined in claim 1; Rbis asdefined in claim 1.
12. 12. The following compounds, an isomer thereof or apharmaceutically acceptable salt thereof, 0 0a 0 HN\S NH HN\ S NH N N ~.- HN:\S N- HHN N-. N N N N NH

    0 0 0 0 HNbNSN:H' N I- \ I NI HN\ S- NH HNN HN- \N N N N _ N N- NN

    0 0 0 0 HN\S H HN\ IN HN NH NH: N N N N N NX N Z NH NN.

    0 0 0 0 HN\ S NH HN\ S NH HN S NH S N N N'

    NH N
13. 13. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 12, selected from the group consisting of 0 o 0 0 HN\S] HN H N S HHN N H ~ ~ ~NN \' N NHH N

    0 0 0 0 HN S NH HN\ S NH H)N HH HN N\ I N' N N- \ NA rHNt \elb&

    N~ S NH H 'NN

    0 0 0 0 HN\W SNH 1 HN\W NH HN\ S INH HN\ S NNH NN/ N- )"6N N-N - \ 4

    0 0 0 0 HN \ S NH HN \ S NH HN\ S NH HN S NH HN \ " N NH NN NH N NN

    0 0 0 0 HN \ S 'NH HN \ S NH H \ S N H N\ S NH N- \ NH N H2
14. 14. A pharmaceutical composition comprising a therapeutically effective amount of the compound or the pharmaceutically acceptable salt thereof according to any one of claims 1-13 and a pharmaceutically acceptable carrier.
15. 15. The compound or the pharmaceutically acceptable salt thereof according to any one of claims 1-13 or the pharmaceutical composition according to claim 14 for use in treating a Cdc7 kinase-mediated disease, wherein optionally, the Cdc7 kinase-mediated disease is selected from a tumor; optionally the Cdc7 kinase-mediated disease is selected from colorectal cancer and pancreatic cancer.

    Example 1-1 Example 1-1 Example 1-1 tumor volume

    Days after administration Figure 1

    Group 1

    Group 2

    Group 3

    Group 4

    Group 5

    Figure 2